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在蛋白质免疫印迹法中通过定性光学方法检测转膜效率和上样量是否均等。

Checking transfer efficiency and equal loading via qualitative optical way in western blotting.

作者信息

Gong Jun-Hua, Gong Jian-Ping, Zheng Kai-Wen

机构信息

Department of Hepatobiliary Surgery, Second Hospital Affiliated to Chongqing Medical University, Chongqing, P. R. China.

出版信息

Electrophoresis. 2017 Nov;38(21):2786-2790. doi: 10.1002/elps.201700266. Epub 2017 Aug 14.

Abstract

The ability to determine that successful transfer and equal loading occur prior to using primary antibodies is important. And total protein staining is commonly used to check transfer efficiency and normalization, which play a crucial role in western blotting. Ponceau S and coomassie blue are commonly used, but there are disadvantages reported in recent years. Therefore, we are interested in finding another method, which is cheap, easy and fast. As we know, protein binding region of PVDF membrane is still hydrophilic when carbinol volatilizes, however, the non-protein binding region of PVDF membrane became hydrophobic again. And this different wettability between non-protein binding region and protein binding region of Polyvinylidene difluoride membrane may be used to check transfer efficiency and equal loading in western blotting. Based on the principle above, we describe an optical approach where an experimenter can observe that the proteins have been transferred to the membrane without any staining within minutes.

摘要

在使用一抗之前确定成功转印和等量上样的能力很重要。总蛋白染色常用于检查转印效率和标准化,这在蛋白质印迹法中起着关键作用。丽春红S和考马斯亮蓝是常用的,但近年来有报道称它们存在缺点。因此,我们有兴趣寻找另一种便宜、简便且快速的方法。我们知道,当甲醇挥发时,聚偏二氟乙烯(PVDF)膜的蛋白质结合区域仍然是亲水的,然而,PVDF膜的非蛋白质结合区域又变回疏水的了。聚偏二氟乙烯膜的非蛋白质结合区域和蛋白质结合区域之间这种不同的润湿性可用于检查蛋白质印迹法中的转印效率和等量上样。基于上述原理,我们描述了一种光学方法,实验者可以在几分钟内观察到蛋白质已在无任何染色的情况下转移到膜上。

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